Autopsies of soldiers killed by mustard gas in World War I indicated that sulfur mustard has a disproportionate effect on rapidly dividing cells and suggested that sulfur mustard compounds might have antitumor effects. Indeed, early researchers attempted to treat cancer by direct injection of sulfur mustard into tumors. This research was limited by the extreme toxicity of sulfur mustard compounds and nitrogen mustard analogs, such as mechlorethamine, were investigated as less toxic alternatives.

Because of the lack of selectivity of most mechlorethamine analogs, prodrugs, such as phosphoramide compounds, which can be activated by the high concentration of phosphoramidases present in neoplastic cells, have been investigated. Two phosphoramide alkylating agents, cyclophosphamide (CPA) and the isomeric compound ifosfamide (IFOS), have demonstrated effectiveness in the treatment of a broad range of solid tumors and hematological cancers (Zhang et al., Current Drug Therapy 1: 55-84 (2006)). CPA and IFOS are used both as single agents as well as in combination with other anticancer agents to obtain synergistic antitumor effects. In addition to its application in cancer, CPA can also be used as an immunosuppressant to treat autoimmune diseases such as rheumatoid arthritis and systemic lupus erythematosus (SLE) (Petri et al., Lupus 13:366-371 (2006); Leandro et al. Ann., Rheum. Dis. 61: 883-888 (2002); Verberg et al., Arthritis Rheum. 52: 421-424 (2005)).

The metabolism of CPA and IFOS has been described in detail by Zhang et al. (Zhang et al., Current Drug Therapy 1: 55-84 (2006)). CPA and IFOS are prodrugs that are activated intracellularly by 4-hydroxylation by the cytochrome (CYP) P450 oxidases, primarily CYP3A4, CYP2C9 and CYP2B6 in the liver, to produce cytotoxic nitrogen mustards that can react with DNA. Acrolein is a byproduct of this reaction. The in vivo metabolism of CPA and IFOS also involves inactivation by N-decholoroethylation by CYP3A4/5 and CYP2B6 prior to their conversion to the nitrogen mustards, resulting in production of dechloroethylated metabolites and the byproduct chloroacetaldehyde (CAA). Acrolein and CAA are implicated in toxicities of CPA and IFOS that are unrelated to the cytotoxic mechanism of action of the nitrogen mustard molecules (Zhang et al., Current Drug Therapy 1: 55-84 (2006)). Acrolein causes the urotoxicity, hemorrahagic cystitis, and liver damage, and CAA causes neurotoxicity and has also been implicated in renal toxicity. Co-administration of the sulfhydryl compounds, mesna and amifostine, which react specifically with acrolein in the urinary tract, can reduce the urotoxicity of acrolein but does not eliminate other toxicities (Zaki et al., Toxicol. In Vitro 17: 397-402 (2003)).
The nitrogen mustards of CPA and IFOS, phosphoramide mustard and isophosphoramide mustard, are bifunctional alkylating agents that bind covalently to nucleophilic groups of nucleic acids. At pH≧7, the mustards are dechlorinated to produce carbonium ions that react covalently with N7 of guanine residues. The reaction is referred to as DNA alkylation. Both inter- or intra-strand crosslinks result from the ability of each mustard molecule to react with two guanine residues (Zhang et al., Current Drug Therapy 1: 55-84 (2006)). Because the inter-strand crosslinks prevent strand separation required for DNA replication, DNA-alkylation is considered to be the major mechanism responsible for the inhibition of cell division by CPA and IFOS. In addition to the antiproliferative (cytostatic) effect, the DNA damage also induces apoptosis, i.e., programmed cell death (O'Conner et al., Cancer Res. 1: 6550-6557; (1991); Bahtia et al., Clin. Cancer Res. 1: 873-880 (1995)). The cytotoxic/cytostatic effects of the nitrogen mustards are mainly responsible for the antitumor activity of CPA and IFOS and, by preventing the proliferative expansion of autoreactive lymphocytes, also for the immunosuppressant activity of CPA in autoimmune disease. However, cross-linking of DNA in normal tissues by the nitrogen mustards also causes cytotoxic, mechanism-based collateral damage, particularly myelosuppression resulting in leucocytopenia, which is the principal dose-limiting hematological toxicity (Zhang et al., Current Drug Therapy 1: 55-84 (2006)).
Although phosphoramide mustard and isophosphoramide are chemically similar, isophosphoramide mustard interacts with DNA with a higher affinity than phosphoramide mustard (Boal et al., J. Med. Chem. 32: 1768-1773; 1989). Structural differences involving the intramolecular distance between the chloroethyl groups and their orientation appear to be responsible for the different affinities of the two mustards (Springer et al., J. Org. Chem. 63: 7218-7222 (1998)).
By administering cytotoxic nitrogen mustards directly to cancer patients, the “off-target” toxicities and the drug resistance associated with the prodrugs may be reduced. IPM has been synthesized and preliminary biological evaluations of the compound have been conducted; but, unfortunately, IPM itself is unstable and difficult to use directly for human treatment. Stabilized formulations of IPM might further reduce toxicity and allow metronomic administration of doses that are sufficient for both direct cytotoxicity against the tumor and antiangiogenic activity. Improved methods for formulation and manufacture of IPM and analogues and salts thereof are needed.